Project Summary PepT1 is mainly expressed in brush-border membranes of enterocytes in the small intestine. PepT1 has a differential pattern of expression in the small intestine. Along the vertical axis, PepT1 is most abundant at the villous tip, with expression decreasing towards the crypt, whereas along the longitudinal axis, the PepT1 level increases from the duodenum to the ileum. However, PepT1 mRNA or protein is expressed at very low levels or is not expressed in the colon. Consistent with these findings, hPepT1 is expressed in the small intestine-like cell line Caco2-BBE but not in the colonic-like cell line HT29-Cl.19A. Although PepT1 is not normally expressed in colonic epithelial cells, we and others have reported that hPepT1 is expressed in epithelial cells of the chronically inflamed colon and in the colonic mucosa of patients with shortbowel syndrome, confirming that hPepT1 expression is induced in colonic epithelial cells under pathological conditions. Recently, we have demonstrated that hPepT1 transports the antiinflammatory tripeptide KPV that has been shown to reduce intestinal inflammation. Our main hypothesis is that regulated PepT1 expression along the crypt[unreadable]villus axis is required for its physiological role in nutrient absorption, whereas aberrant expression in the colon under pathological conditions modulates disease manifestation. The first aim of our proposal will investigate the regulatory role of miR-619 in PepT1 differential expression along the crypt[unreadable]villus axis and the sequence target of miR-619 on the PepT1 gene. Second, we aim to determine the roles of the transcription factor Cdx2 and miR-619 in determining the levels of colonic PepT1 expression and will assess its effects on colonic epithelial functions in the third aim. It is important to understand the regulation of PepT1 expression in the small intestine because PepT1 controls overall protein absorption via the PepT1-mediated di/tripeptide transport that is essential for individuals to remain healthy. In addition, the development of therapeutic strategies capable of controlling PepT1 expression could be important for enteral nutrition and PepT1-mediated drug delivery. Finally, it is also important to understand the transcriptional and post-transcriptional mechanisms by which PepT1 can be expressed in colonic epithelial cells and the pathophysiological consequences of this expression. The ultimate goal is to identify and develop therapeutic strategies to control abnormal PepT1 expression under pathological conditions.